Limonene and perillyl alcohol are monoterpenes that are dietary components as well as food flavoring agents and scent additives. Limonene is abundant in orange peel oil, and perillyl alcohol in cherry and spearmint. These compounds have shown potential as natural anti-cancer agents. The monoterpenes have been proposed to inhibit oncogenic protein prenylation by blocking the activity of both farnesyltransferase (FTase) and geranylgeranyltransferase type I (GGTase-I). Combination treatment with synthetic inhibitors of FTase and GGTase-I has been pursued as an avenue of cancer therapy, but many of available GGTase-I inhibitors are excessively toxic. The monoterpenes are naturally occurring, nontoxic inhibitors of both prenyltransferases. These studies seek to elucidate the structural basis of this dual inhibitory effect of the monoterpenes. The working hypothesis of this proposal is that the monoterpenes interfere with substrate binding to FTase and GGTase-I by occupying the similar hydrophobic binding pockets of the prenyltransferases. A combination of x-ray crystallography, mass spectrometry, and biochemical analysis will be used to complete two specific aims: 1. Determine the structural changes of FTase and GGTase-I that are induced by phosphorylation and establish how phosphorylation regulates prenylation activity 2. Define the mechanism by which the monoterpenes inhibit FTase/GGTase-I prenylation activity These findings will provide a structural basis for understanding how the monoterpenes suppress tumors via FTase/GGTase-I inhibition. The monoterpene-bound structures of these prenyltransferases will also advance the design of compounds that are more effective than the monoterpenes but less toxic than the existing synthetic inhibitors. Such dual inhibitors of FTase and GGTase-I would be useful for treatment of various human malignancies.